NaN results on CLC GW

[conejeros]

Hi Guys, I´m really newby on this so I appreciate the help a lot.
Recently I sent samples for RNAseq to Macrogen and I got the results, including a basic bioinformatic analisis. Now, I´m trying to repeat what they did from scratch in order to learn about it and being able to apply further analysis, on CLC GWB software. However I cant get close to what Macrogen got.
Macrogen results included a number of high RPKM values, when using a EST library as reference sample. However, when I perform the basic RNAseq analisys towards the same EST library as a reference, on the CLC GWB, I only get NaN for expression and RPKM.
Can anyone help me with this, tell me what I´m missing, may be recomend me some basic reading?
Thanks a lot.
Cheers!

[GenoMax]

Did you do any QC/trimming of your data before you tried to do the alignments? In what format is your "reference" transcriptome (one transcript per gene as CLC specifies)?

Unless Macrogen used CLC to do the analysis it may be difficult to "reproduce" what they did using CLC. Every step (QC, trimming, alignment, counting) they did is not going to map well in CLC since CLC uses many proprietary algorithms. At the best you can hope to get a (very) similar DE gene list (but the fpkm numbers may not match).

Having said that have you contacted CLC tech support about this set of questions? If you paid for a license you may as well get support from them. They may be able to remote into your machine and look at your analysis directly.

[conejeros]

Thank you very much for your help. I understand the numbers wont be the same, but I´m intending to do something at least similar.
I haven´t done anything with the raw data (as I said, sorry, newby here). I´ll try repeating all their processes with the CLC equivalent, and let know here,
thanks again!

[GenoMax]

Ah. I thought you had done an analysis with CLC and were not able to reproduce the result. Do the best approximation of what they did (ask them for what programs they used and the settings for those). That should help you locally with CLC options.